Purification And Characterization Of Alpha-L-Arabinofuranosidases From Geobacillus Stearothermophilus Strain 12
| Autor: | Kadriye Inan Bektas, Ali Osman Belduz, Sabriye Canakci, Ali Sevim, Elif Sevim, Iclal Sahin |
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| Přispěvatelé: | Kırşehir Ahi Evran Üniversitesi, Mühendislik-Mimarlık Fakültesi, Genetik ve Biyomühendislik Bölümü |
| Rok vydání: | 2017 |
| Předmět: |
0106 biological sciences
0301 basic medicine Stereochemistry thermophilic bacteria thermostable enzymes Plant Science alpha-L-Arabinofuranosidases 01 natural sciences Biochemistry law.invention Geobacillus stearothermophilus 03 medical and health sciences law 010608 biotechnology Genetics Glycoside hydrolase Molecular Biology Ecology Evolution Behavior and Systematics chemistry.chemical_classification Expression vector Strain (chemistry) Thermophile Substrate (chemistry) Cell Biology 030104 developmental biology Enzyme chemistry Recombinant DNA Animal Science and Zoology |
| Popis: | WOS: 000410907000002 In order to characterize two alpha-L-arabinofuranosidases (alpha-L-AFases), Abf1Geo12 and Abf2Geo12, produced by Geobacillus stearothermophilus strain 12, the genes (abf 1 and abf 2) coding for these enzymes were cloned and sequenced. Based on the protein sequence similarities, approximately 57 kDa two alpha-L-AFases were assigned to the glycoside hydrolase family 51. To obtain pure enzymes, the abf 1 and abf 2 genes were cloned into pET28a+ expression vector and recombinant alpha-L-AFases were produced in E. coli BL21(DE3): pLysS. Characterization of recombinant alpha-L-AFases revealed that Abf1Geo12 and Abf2Geo12 were active in a broad temperature range from 50 to 85 degrees C and from 40 to 80 degrees C, respectively. Also, the Abf1Geo12 was active in a broad pH range from 5.0 to 9.0. The optimum pH and temperature for Abf1Geo12 were determined as pH 6.0 and 65 degrees C, respectively, whereas the optimum pH and temperature for Abf2Geo12 were determined as pH 5.5 and 60 degrees C, respectively. Based on characterization studies, it was determined that the Abf1Geo12 was more stable than Abf2Geo12 and previously identified alpha-L-AFases from G. stearothermophilus. Using p-nitrophenyl alpha-L-arabinofuranoside as a substrate, the Km and Vmax values for Abf1Geo12 and Abf2Geo12 were determined as 0.31 mM and 290 U/mg for the former enzyme and 0.19 mM and 213.2 U/mg for the latter enzyme, respectively. The activities of Abf1Geo12 and Abf2Geo12 were strongly inhibited by 1 mM Hg2+. Interestingly, Cu2+ and Co2+ stimulated the activity of Abf1Geo12, but they reduced the activity of Abf2Geo12. The recombinant enzymes released L-arabinose from sugar beet arabinan, arabinobiose, arabinotriose, arabinotetraose and arabinopentaose. Consequently, these characterized two enzymes may be used in industrial fields since they are stable at high temperatures. Scientific and Research Council of Turkey (TUBITAK)Turkiye Bilimsel ve Teknolojik Arastirma Kurumu (TUBITAK) [211T116] This work was supported by The Scientific and Research Council of Turkey (TUBITAK, Grant No.: 211T116). |
| Databáze: | OpenAIRE |
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